title: A novel protease activity assay method based on an engineered autoinhibited protein using an enzyme-linked immunoassay.
abstract: Proteases are involved in various biological phenomena, and their aberrant activity can be an important indicator of disease. Thus, various methods have been developed to analyze the activities of proteases, but their wide application has been hampered because each method has drawbacks. In this report, we propose a new protease assay method based on an engineered autoinhibited protein and enzyme-linked immunoassay (ELISA) in which a protease of interest activates the autoinhibited protein and the signal is amplified via ELISA. Using this concept a sensitive assay method for MMP2 and caspase-3 was developed. The limit of detection for the two proteases was as low as 7 pM for MMP2 and 0.1 pM for caspase-3. The autoinhibited protein is designed modularly, and the new platform is general enough for the development of assay methods for other proteases with minimal modification.
authors.affiliation: Department of Molecular Science and Technology, Ajou University, Suwon, 443-749, Korea. taehyeonyoo@ajou.ac.kr.
authors.forename: Hyun Kyung
authors.initials: HK
authors.lastname: Yoon
authors.forename: Tae Hyeon
authors.initials: TH
authors.lastname: Yoo
mesh_terms: D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D010447:Peptide Hydrolases / Q000378:metabolism*; D015202:Protein Engineering
chemical_list: D010447:Peptide Hydrolases